scholarly journals The expression of nicotinamide N-methyltransferase increases ATP synthesis and protects SH-SY5Y neuroblastoma cells against the toxicity of Complex I inhibitors

2011 ◽  
Vol 436 (1) ◽  
pp. 145-155 ◽  
Author(s):  
Richard B. Parsons ◽  
Shylesh Aravindan ◽  
Anusha Kadampeswaran ◽  
Emily A. Evans ◽  
Kanwaljeet K. Sandhu ◽  
...  
Keyword(s):  
Cell Line ◽  
Complex I ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cells ◽  
Atp Synthesis ◽  
Neuroblastoma Cell Line ◽  
Cellular Effects ◽  
Endogenous Expression ◽  
Complex I Activity

NNMT (nicotinamide N-methyltransferase, E.C. 2.1.1.1) catalyses the N-methylation of nicotinamide to 1-methylnicotinamide. NNMT expression is significantly elevated in a number of cancers, and we have previously demonstrated that NNMT expression is significantly increased in the brains of patients who have died of Parkinson's disease. To investigate the cellular effects of NNMT overexpression, we overexpressed NNMT in the SH-SY5Y cell line, a tumour-derived human dopaminergic neuroblastoma cell line with no endogenous expression of NNMT. NNMT expression significantly decreased SH-SY5Y cell death, which correlated with increased intracellular ATP content, ATP/ADP ratio and Complex I activity, and a reduction in the degradation of the NDUFS3 [NADH dehydrogenase (ubiquinone) iron–sulfur protein 3] subunit of Complex I. These effects were replicated by incubation of SH-SY5Y cells with 1-methylnicotinamide, suggesting that 1-methylnicotinamide mediates the cellular effects of NNMT. Both NNMT expression and 1-methylnicotinamide protected SH-SY5Y cells from the toxicity of the Complex I inhibitors MPP+ (1-methyl-4-phenylpyridinium ion) and rotenone by reversing their effects upon ATP synthesis, the ATP/ADP ratio, Complex I activity and the NDUFS3 subunit. The results of the present study raise the possibility that the increase in NNMT expression that we observed in vivo may be a stress response of the cell to the underlying pathogenic process. Furthermore, the results of the present study also raise the possibility of using inhibitors of NNMT for the treatment of cancer.

TTX-Sensitive and -Resistant Na+ Currents, and mRNA for the TTX-Resistant rH1 Channel, Are Expressed in B104 Neuroblastoma Cells

1997 ◽  
Vol 77 (1) ◽  
pp. 236-246 ◽  
Author(s):  
Xiang Q. Gu ◽  
Sulayman Dib-Hajj ◽  
Marco A. Rizzo ◽  
Stephen G. Waxman
Keyword(s):  
Steady State ◽  
Cell Line ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cells ◽  
Neuroblastoma Cell Line ◽  
Membrane Excitability ◽  
Rt Pcr ◽  
Α Subunit ◽  
Time To Peak ◽  
Steady State Inactivation

Gu, Xiang Q., Sulayman Dib-Hajj, Marco A. Rizzo, and Stephen G. Waxman. TTX-sensitive and -resistant Na+ currents, and mRNA for the TTX-resistant rH1 channel, are expressed in B104 neuroblastoma cells. J. Neurophysiol. 77: 236–246, 1997. To examine the molecular basis for membrane excitability in a neuroblastoma cell line, we used whole cell patch-clamp methods and reverse transcription-polymerase chain reaction (RT-PCR) to study Na+ currents and channels in B104 cells. We distinguished Tetrodotoxin (TTX)-sensitive and -resistant Na+ currents and detected the mRNA for the cardiac rH1 channel in B104 cells. Na+ currents could be recorded in 65% of cells. In the absence of TTX, mean peak Na+ current density was 126 ± 19 pA/pF, corresponding to a channel density of 2.7 ± 0.4/μ2 (mean ± SE). Time-to-peak (t-peak), activation (τm), and inactivation time constants (τh) for Na+ currents in B104 cells were 1.0 ± 0.04, 0.4 ± 0.06, and0.9 ± 0.04 ms at −10 mV. The peak conductance-voltage relationship had a V 1/2 of −39.8 ± 1.5 mV. V 1/2 for steady-state inactivation was −81.6 ± 1.5 mV. TTX-sensitive and -resistant components of the Na current had half-maximal inhibitions (IC50), respectively, of 1.2 nM and, minimally, 575.5 nM. The TTX-sensitive and-resistant Na+ currents were kinetically distinct; time-to-peak, τm, and τh for TTX-sensitive currents were shorter than for TTX-resistant currents. Steady-state voltage dependence of the two currents was indistinguishable. The presence of TTX-sensitive and-resistant Na+ currents, which are pharmacologically and kinetically distinct, led us to search for mRNAs known to be associated with TTX-resistant channels, in addition to the α subunit mRNAs, which have previously been shown to be expressed in these cells. Using RT-PCR and restriction enzyme mapping, we were unable to detect αSNS, but detected mRNA for rH1, which is known to encode a TTX-resistant channel, in B104 cells. B104 neuroblastoma cells thus express TTX-sensitive and -resistant Na+ currents. These appear to be encoded by neuronal-type and cardiac Na+ channel mRNAs including the RH1 transcript. This cell line may be useful for studies on the rH1 channel, which is known to be mutated in the long-QT syndrome.

scholarly journals Inhibition of the SK-N-MC human neuroblastoma cell line in vivo and in vitro by a novel nutrient mixture

2013 ◽  
Vol 29 (5) ◽  
pp. 1714-1720 ◽  
Author(s):  
M. WAHEED ROOMI ◽  
TATIANA KALINOVSKY ◽  
NUSRATH W. ROOMI ◽  
ALEKSANDRA NIEDZWIECKI ◽  
MATTHIAS RATH
Keyword(s):  
Cell Line ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cell Line ◽  
Human Neuroblastoma Cell ◽  
Human Neuroblastoma ◽  
Human Neuroblastoma Cell Line

scholarly journals Camptothecin Regulates Microglia Polarization and Exerts Neuroprotective Effects via Activating AKT/Nrf2/HO-1 and Inhibiting NF-κB Pathways In Vivo and In Vitro

2021 ◽  
Vol 12 ◽  
Author(s):  
Dewei He ◽  
Shoupeng Fu ◽  
Ang Zhou ◽  
Yingchun Su ◽  
Xiyu Gao ◽  
...  
Keyword(s):  
Cell Line ◽  
Inflammatory Responses ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cell Line ◽  
Mechanism Study ◽  
Antitumor Effects ◽  
Neuroprotective Effects ◽  
Microglia Polarization

Microglia, the main immune cells in the brain, participate in the innate immune response in the central nervous system (CNS). Studies have shown that microglia can be polarized into pro-inflammatory M1 and anti-inflammatory M2 phenotypes. Accumulated evidence suggests that over-activated M1 microglia release pro-inflammatory mediators that damage neurons and lead to Parkinson’s disease (PD). In contrast, M2 microglia release neuroprotective factors and exert the effects of neuroprotection. Camptothecin (CPT), an extract of the plant Camptotheca acuminate, has been reported to have anti-inflammation and antitumor effects. However, the effect of CPT on microglia polarization and microglia-mediated inflammation responses has not been reported. In our study we found that CPT improved motor performance of mice and reduced the loss of neurons in the substantia nigra (SN) of the midbrain in LPS-injected mice. In the mechanism study, we found that CPT inhibited M1 polarization of microglia and promotes M2 polarization via the AKT/Nrf2/HO-1 and NF-κB signals. Furthermore, CPT protected the neuroblastoma cell line SH-SY5Y and dopaminergic neuron cell line MN9D from damage mediated by microglia activation. In conclusion, our results demonstrate that CPT regulates the microglia polarization phenotype via activating AKT/Nrf2/HO-1 and inhibiting NF-κB pathways, inhibits neuro-inflammatory responses, and exerts neuroprotective effects in vivo and in vitro.

scholarly journals Silencing of AURKA augments the antitumor efficacy of the AURKA inhibitor MLN8237 on neuroblastoma cells

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Yan Yang ◽  
Lili Ding ◽  
Qi Zhou ◽  
Li Fen ◽  
Yuhua Cao ◽  
...  
Keyword(s):  
Small Interfering Rna ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cells ◽  
Inhibitory Effect ◽  
Neuroblastoma Cell Line ◽  
Inhibitory Effects ◽  
Rna Transfection ◽  
Interfering Rna

Abstract Background Aurora kinase A (AURKA) has been implicated in the regulation of cell cycle progression, mitosis and a key number of oncogenic signaling pathways in various malignancies including neuroblastoma. Small molecule inhibitors of AURKA have shown potential, but still not as good as expected effects in clinical trials. Little is known about this underlying mechanism. Here, we evaluated the inhibitory effects of AURKA inhibitor MLN8237 on neuroblastoma cells to understand the potential mechanisms responsible for tumor therapy. Methods MLN8237 treatment on neuroblastoma cell line IMR32 was done and in vivo inhibitory effects were investigated using tumor xenograft model. Cellular senescence was evaluated by senescence-associated β-gal Staining assay. Flow cytometry was used to tested cell cycle arrest and cell apoptosis. Senescence-associated signal pathways were detected by western blot. CD133 microbeads and microsphere formation were used to separate and enrich CD133+ cells. AURKA small interfering RNA transfection was carried to downregulate AURKA level. Finally, the combination of MLN8237 treatment with AURKA small interfering RNA transfection were adopted to evaluate the inhibitory effect on neuroblastoma cells. Results We demonstrate that MLN8237, an inhibitor of AURKA, induces the neuroblastoma cell line IMR32 into cellular senescence and G2/M cell phase arrest. Inactivation of AURKA results in MYCN destabilization and inhibits cell growth in vitro and in a mouse model. Although MLN8237 inhibits AURKA kinase activity, it has almost no inhibitory effect on the AURKA protein level. By contrast, MLN8237 treatment leads to abnormal high expression of AURKA in vitro and in vivo. Knockdown of AURKA reduces cell survival. The combination of MLN8237 with AURKA small interfering RNA results in more profound inhibitory effects on neuroblastoma cell growth. Moreover, MLN8237 treatment followed by AURKA siRNA forces senescent cells into apoptosis via suppression of the Akt/Stat3 pathway. Conclusions The effect of AURKA-targeted inhibition of tumor growth plays roles in both the inactivation of AURKA activity and the decrease in the AURKA protein expression level.

Excision of N-myc from chromosome 2 in human neuroblastoma cells containing amplified N-myc sequences

1990 ◽  
Vol 10 (2) ◽  
pp. 823-829
Author(s):  
J D Hunt ◽  
M Valentine ◽  
A Tereba
Keyword(s):  
Neuroblastoma Cell ◽  
Neuroblastoma Cells ◽  
Initial Step ◽  
Neuroblastoma Cell Line ◽  
Chromosome 2 ◽  
Human Neuroblastoma ◽  
Dominant Feature ◽  
The Many ◽  
Tumor Types

Amplification of one of three growth-stimulating myc genes is a common method by which many tumor types gain a proliferative advantage. In metastatic human neuroblastoma, the amplification of the N-myc locus, located on chromosome 2, is a dominant feature of this usually fatal pediatric cancer. Of the many models proposed to explain this amplification, all incorporate as the initial step either disproportionate overreplication of the chromosomal site or recombination across a loop structure. The original locus is retained within the chromosome in the overreplication models but is excised in the recombination models. To test these models, we have used somatic cell hybrids to separate and analyze the chromosomes 2 from a neuroblastoma cell line containing in vivo amplified N-myc. Our results demonstrate that N-myc is excised from one of the chromosomes, suggesting that deletion is a requisite part of gene amplification in a naturally occurring system.

scholarly journals Differentiation-induced Colocalization of the KH-type Splicing Regulatory Protein with Polypyrimidine Tract Binding Protein and the c-srcPre-mRNA

2004 ◽  
Vol 15 (2) ◽  
pp. 774-786 ◽  
Author(s):  
Megan P. Hall ◽  
Sui Huang ◽  
Douglas L. Black
Keyword(s):  
Rna Binding ◽  
Binding Protein ◽  
Neuroblastoma Cell ◽  
Regulatory Protein ◽  
Neuroblastoma Cells ◽  
Neuroblastoma Cell Line ◽  
Polypyrimidine Tract ◽  
Polypyrimidine Tract Binding ◽  
Polypyrimidine Tract Binding Protein

We have examined the subcellular localization of the KH-type splicing regulatory protein (KSRP). KSRP is a multidomain RNA-binding protein implicated in a variety of cellular processes, including splicing in the nucleus and mRNA localization in the cytoplasm. We find that KSRP is primarily nuclear with a localization pattern that most closely resembles that of polypyrimidine tract binding protein (PTB). Colocalization experiments of KSRP with PTB in a mouse neuroblastoma cell line determined that both proteins are present in the perinucleolar compartment (PNC), as well as in other nuclear enrichments. In contrast, HeLa cells do not show prominent KSRP staining in the PNC, even though PTB labeling identified the PNC in these cells. Because both PTB and KSRP interact with the c-src transcript to affect N1 exon splicing, we examined the localization of the c-src pre-mRNA by fluorescence in situ hybridization. The src transcript is present in specific foci within the nucleus that are presumably sites of src transcription but are not generally perinucleolar. In normally cultured neuroblastoma cells, these src RNA foci contain PTB, but little KSRP. However, upon induced neuronal differentiation of these cells, KSRP occurs in the same foci with src RNA. PTB localization remains unaffected. This differentiation-induced localization of KSRP with src RNA correlates with an increase in src exon N1 inclusion. These results indicate that PTB and KSRP do indeed interact with the c-src transcript in vivo, and that these associations change with the differentiated state of the cell.

On the Application of Cultured Neuronal Cell Lines in Neurotoxicological Studies: Implications of Acrylamide-induced Neurite Disintegration

1983 ◽  
Vol 11 (4) ◽  
pp. 194-203
Author(s):  
Elisabet Nyberg ◽  
Erik Walum
Keyword(s):  
Cell Line ◽  
Neuroblastoma Cell ◽  
Membrane Integrity ◽  
Neuronal Cell ◽  
Specific Effect ◽  
Glycolytic Enzyme ◽  
Neuroblastoma Cell Line ◽  
Experimental Conditions ◽  
Neuronal Cell Lines

Summary Cultures of the mouse neuroblastoma cell line C1300, clone N1E115 were exposed to acrylamide at 3.5 x 10-4M for 14 days (subacute situation) or at 2.8 x 10-3 M for 24 hr (acute situation). In the subacute situation the total uptake of 2-deoxy-D-glucose was stimulated. This could be explained by an increase in both the non-specific diffusion and the specific transport. The activity of the glycolytic enzyme, enolase (EC4.2.1.11), was unaffected by exposure to acrylamide, whereas the activity of glyceraldehyde-3-phosphate-dehydrogenase (EC1.2.1.12) was inhibited. Acrylamide had a marked stimulating effect on the respiratory activity of the cells, whereas the incorporation of tritiated leucine remained unchanged. Furthermore, membrane integrity was maintained throughout the acrylamide exposure as judged by an unchanged rate of 2-deoxy-D-glucose-6-phosphate efflux. Corresponding results were obtained in the acute situation. In N1E115 cultures and under the experimental conditions used in this work acrylamide caused neurite degeneration resembling distal axonopathy in vivo. It is suggested that these degenerative changes are not due to a general intoxication of the cells, but rather to a specific effect. Consequently, the N1E115 cell line might be useful in studies of chemically-induced axonopathies.

scholarly journals V max. activation of pp60c-src tyrosine kinase from neuroblastoma neuro-2A

1987 ◽  
Vol 248 (3) ◽  
pp. 691-696 ◽  
Author(s):  
W T Matten ◽  
P F Maness
Keyword(s):  
Cell Line ◽  
Specific Activity ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cells ◽  
Src Kinase ◽  
Maximum Velocity ◽  
Neuroblastoma Cell Line ◽  
Angiotensin I ◽  
Src Tyrosine Kinase ◽  
Km Value

A kinetic analysis of the tyrosine-specific protein kinase of pp60c-src from the C1300 mouse neuroblastoma cell line Neuro-2A and pp60c-src expressed in fibroblasts was carried out to determine the nature of the increased specific activity of the neuroblastoma enzyme. In immune-complex kinase assays with ATP-Mn2+ and the tyrosine-containing peptide angiotensin I as phosphoacceptor substrate, pp60c-src from the neuroblastoma cell line was characterized by a maximum velocity (Vmax.) that was 7-15-fold greater than the Vmax. of pp60c-src from fibroblasts. The neuroblastoma enzyme exhibited Km values for ATP (16 +/- 3 microM) and angiotensin I (6.8 +/- 2.6 mM) that were similar to Km values for ATP (25 +/- 3 microM) and angiotensin I (6.5 +/- 1.7 mM) of pp60c-src from fibroblasts. pp60v-src expressed in Rous-sarcoma-virus-transformed cells exhibited an ATP Km value (25 +/- 4 microM) and an angiotensin I Km value (6.6 +/- 0.5 mM) that approximated the values determined for pp60c-src in neuroblastoma cells and fibroblasts. These results indicate that the pp60c-src kinase from neuroblastoma cells has a higher turnover number than pp60c-src kinase from fibroblasts, and that the neural form of the enzyme would be expected to exhibit increased catalytic activity at the saturating concentrations of ATP that are found intracellularly.

Sign in / Sign up

Export Citation Format

Share Document